Characterisation of the 33kDa piroplasm surface antigen of Theileria orientalis/sergenti/buffeli isolates from West Java, Indonesia.

The immunodominant 33/35kDa antigen of a Theileria isolate from West Java, Indonesia, was characterised and immuno-affinity purified by use of a monoclonal antibody, KUL-a4, and was shown to be representative of the T. orientalis/sergenti/buffeli group. The aminoterminal sequence of the purified 35kDa peptide (20 residues) was determined by automated Edman degradation and found to correspond to the predicted amino acid sequence of a prospective p33 gene previously sequenced from the same isolate. The cleavage site of a putative signal peptide was identified and conforms the (-3, -1) rule for signal peptidases. The existence of dimeric and trimeric forms of the p33/35 antigen is hypothesised from Western blot profiles. KUL-a4 appeared specific for the T. orientalis/sergenti/buffeli group. It did not recognise in indirect fluorescence antibody test (IFAT), intraerythrocytic bodies of Anaplasma marginale or piroplasms and schizonts of T. mutans, T. parva and T. annulata, whereas cattle antisera raised to these species showed cross-reactivity in IFAT. It however, appeared weakly cross-reactive in Western blot and ELISA, with the 34kDa piroplasm antigen of one T. annulata (Gharb) isolate. The present study indicates that the isolated antigen belongs to the p33/34 antigen family described within the T. sergenti/orientalis/buffeli group, and documents the group-specificity of one of its epitopes.     

Predominance of the papGII allele with high sequence homology to that of human isolates among avian pathogenic Escherichia coli (APEC)

Avian pathogenic Escherichia coli (APEC) are often found in poultry and are responsible for a set of diseases, commonly referred to as avian colibacillosis. One of the important virulence factors is adhesion to different epithelial surfaces, which is mediated by pili. P pili are thought to play a role by means of their PapG adhesin, which occurs in three molecular variants: PapGI, PapGII and PapGIII. This study is the first to determine and analyse the distribution of the different papG alleles in APEC. Our results show a significant predominance of the papGII allele above all other alleles or allele combinations. No statistically significant associations could be found between papG allele distribution and the type of bird, organ of isolation and O serogroup. Finally, the papGII and papGIII sequences showed high homology with mammalian (including human) source papG sequences.     

F4 receptor-independent priming of the systemic immune system of pigs by low oral doses of F4 fimbriae

Oral administration of F4 fimbriae of Escherichia coli induces intestinal mucosal immune responses in F4 receptor-positive (F4R(+)) pigs, but not in F4R(-) pigs. We examined whether F4 fimbriae in F4R(-) animals behave like a food antigen and can induce oral tolerance. Therefore, F4R(+) and F4R(-) pigs were fed 2mg of F4 and challenged i.m. to evaluate the effect of oral F4 on the systemic immune system. As control antigen, two different oral doses (2 and 600 mg) of OVA were used. Thirty days after the i.m. OVA challenge, the OVA-specific serum IgG titre in 600 mg-fed pigs was lower than that in non-fed animals, indicating that tolerance was induced. Conversely, in the 2mg-fed pigs a rapid increase of OVA-specific IgG with higher titres than those in non-fed pigs was seen following challenge, indicating a priming of the systemic immune system. A similar priming was seen in both F4-fed F4R(-) and F4R(+) pigs. Upon challenge, non-fed pigs displayed a primary immune response with a slow increase of F4-specific serum IgG, whereas F4-fed F4R(-) and F4R(+) pigs showed secondary responses with a rapid increase of serum IgG. This was expected in F4R(+) pigs, as in these animals oral F4 induces F4-specific antibody-secreting cells in the spleen, suggesting a priming of the systemic immune system. However, also the F4-fed F4R(-) pigs displayed a secondary response, despite the failure to detect a response upon oral F4 administration. These findings suggest that the F4 antigen, at a dose of 2 mg, behaves like a common food antigen in F4R(-) pigs, namely it induces a systemic priming.     

Where do we go in protection against pathogens?

A few reflections are made on the future strategies and approaches for combating infectious diseases of livestock, keeping into consideration public concern on food quality and availability. The policy of eradication of pathogens/diseases instead of vaccination has rendered livestock naive to certain pathogens and as a consequence susceptible to epidemic outbreaks, even more so as we have to conclude that outside attack is quite often difficult to control. Therefore, immunoprophylactic measures in livestock remain top priority and will in the future focus more on the induction of mucosal immunity and the activation of innate immunity.     

Enhanced specific antibody response to bovine serum albumin in pigeons due to L-carnitine supplementation

1. Thirty adult female pigeons (Columba livia domestica) were randomly divided into 3 equal groups; the 1st and 2nd groups were immunised with bovine serum albumin (BSA) at 0 and 20 d, the 2nd group also received 1 g L-carnitine per litre of drinking water from -5 to 25 d post-immunisation (dpi) and the 3rd group, a control group, received neither treatment. 2. Body weights and serum samples were taken at 0, 5, 10, 15, 20, 25, 30 and 35 dpi. 3. Both BSA-specific IgG and IgM responses were enhanced by about 10% by L-carnitine supplementation. 4. L-carnitine supplemented pigeons showed a higher water consumption. Body weight loss during the onset of the immune response showed a slight tendency to be counteracted by L-carnitine supplementation. 5. The impact of L-carnitine on resistance and resilience to an immunological challenge is discussed.     

Monoclonal antibodies identify phenotypically and functionally distinct cell types in the bovine lymphoid system

Monoclonal antibodies were produced against bovine lymphoid cells. The reactivities of the antibodies for membrane determinants were examined on both cell suspensions and cryostat tissue sections prepared from bovine blood, thymus, spleen and lymph nodes. The antibodies were putatively grouped into sets which reacted with monomorphic and polymorphic determinants associated with bovine class I and class II major histocompatibility complex (MHC) antigens (MAbs P12 and P3, and R1 and P2 respectively), or associated with differentiation antigens expressed on T cells and monocytes (MAb P5) or exclusively on monocytes (MAb P8). The antibodies were used to identify the surface phenotypes of cells which stimulate (R1+ P5+ P8+) and proliferate (R1- P5+ P8-) in the bovine mixed leukocyte cultures, and cells which proliferate in response to the mitogen, concanavalin A (R1- P5+).     

Development of immunocompetence of broiler chickens

Subpopulations of T-cells, B-cells, macrophages and ellipsoid-associated reticular cells (EARC) could be demonstrated by immunohistochemical staining early in the development of chicken spleen. However, the typical structures of the spleen, such as the peri-arteriolar lymphoid sheath (PALS) and the ellipsoids with their surrounding ring of macrophages, were only formed around embryonic day (ED) 20. These structures and especially the B-cell compartment, i.e., the peri-ellipsoid lymphoid sheath (PELS) gradually matured during the first week posthatch.

Therefore, we analysed at what age broiler chickens could generate a humoral response against the thymus-dependent antigen bovine serum albumin (BSA). Chickens were immunised in ovo (ED16 and ED18) and at 1, 7 and 12 days of age and subsequent BSA-specific immunoglobulin (Ig) M and IgG responses were measured up to 10 days postimmunisation (DPI). No major differences were observed in the relative growth rates, while hatchability was only slightly reduced. Only in chicks immunised on 12 days of age, IgM and IgG responses were high with a normal kinetic pattern. In chicks immunised on 7 days of age, responses were just detectable, but they were absent in chicks immunised in ovo and on the day of hatching (Day 1).

In a subsequent experiment, 1-, 7- and 12-day-old chicks were BSA-immunised and Ig responses were measured for a longer period up to the age of 28 days. The IgG response of chicks immunised at 1 day of age was lower and occurred later (from 28 DPI) than the response of chicks immunised at 7 and 14 days of age (from 14 DPI). It was not increased by a booster immunisation on 29 days of age, in contrast to the response of chicks immunised at 7 and 14 days of age. These findings indicate that vaccination at 1 day of age does not activate the B-cell response resulting in antibody production and support the idea that the immune function of the late embryonic and neonatal chickens is not entirely developed due to the incomplete structural organisation of their secondary immune organs.     

Comparison of immune responses in parenteral FaeG DNA primed pigs boosted orally with F4 protein or reimmunized with the DNA vaccine

We previously showed that an intradermal (i.d.) FaeG DNA prime (2x)-oral F4 protein boost immunization induces a systemic response and weakly primes a mucosal IgG response in pigs, especially when plasmid vectors encoding the A and B subunit of the E. coli thermo-labile enterotoxin (LT) are added to the DNA vaccine. In the present study, we evaluated whether addition of 1alpha,25-dihydroxyvitamin D(3) (vitD(3)) to the DNA vaccine could further enhance this mucosal priming and/or modulate the antibody response towards IgA. To further clarify priming of systemic and mucosal responses by the i.d. DNA vaccination, we firstly compared the localization of the F4-specific antibody response in pigs that were orally boosted with F4 to that in pigs that received a third i.d. DNA immunization and secondly evaluated cytokine mRNA expression profiles after i.d. DNA vaccination. The i.d. DNA prime (2x)-oral F4 boost immunization as well as the 3 i.d. DNA vaccinations induced mainly a systemic response, with a higher response observed following the heterologous protocol. Co-administration of vitD(3), and especially of the LT vectors, enhanced this response. Furthermore, only the heterologous immunization resulted in a weak mucosal priming, which appeared to require the presence of the LT vectors or vitD(3) as adjuvants. In addition, the LT vectors strongly enhanced the FaeG-specific lymphocyte proliferation and this was accompanied by the absence of a clear IL-10 response. However, despite two DNA immunizations in the presence of these adjuvants and an oral F4 boost, we failed to demonstrate the secretory IgA response needed to be protective against enterotoxigenic E. coli.     

Bovine mononuclear phagocytic cells: identification by monoclonal antibodies and analysis of functional properties

Monoclonal antibodies (mAb) which react with bovine monocytes have been produced. These include three mAb (P8, IL-A22 and IL-A24) that recognize the majority of monocytes and granulocytes in peripheral blood; two of these mAb were also shown to react with 30-40% of cells in bone marrow, including both monocytic and granulocytic cells, and with variable percentages of tissue macrophages. Thus these mAb can act as markers for myeloid cells in haemopoietic tissues and for monocytes in cell populations devoid of granulocytes. A further two mAb (IL-A23 and IL-A25) recognize monocytes and/or macrophages. The reactivity of one of these mAb (IL-A25) appears to be mainly restricted to pulmonary macrophages. The other mAb reacts with a variable proportion of blood monocytes and generally with a higher percentage of tissue macrophages, suggesting that its expression may relate to activation or maturation of monocytes. In order to study the functional properties of peripheral blood monocytes, techniques were developed for obtaining populations of peripheral blood mononuclear cells (PBM) depleted of monocytes to less than 0.2% and monocyte populations of greater than 97% purity. Removal of monocytes from PBM abrogated the capacity of the cells to proliferate in response to Con A and PBS, although addition of 2-mercaptoethanol to the cultures restored proliferation. In both allogeneic and autologous mixed leukocyte cultures (MLC), monocytes were required in the stimulator cell populations for induction of the proliferative responses, and both responses could be elicited with purified monocytes. However, proliferation in the autologous MLC occurred only with responder cell populations that were depleted of monocytes. Moreover, it was shown that addition of more than 5% unirradiated monocytes to the autologous MLC suppressed proliferation. These findings indicate that monocytes play an important role in the induction and regulation of cellular immune responses in cattle. Two of the mAb that react with monocytes and granulocytes were tested for their capacity to inhibit proliferative responses of PBM to mitogens, alloantigens or the soluble antigen, KLH.(ABSTRACT TRUNCATED AT 400 WORDS)